One of insulin's major biologic effects is stimulation of glucose transport, and detailed knowledge of the molecular events which mediate this bioeffect will lead to improved understanding of mechanisms of insulin action as well as disease states characterized by insulinresistance. The thiazoladinedione (TZD) class ofinsulin sensitizers binds to the PPARy receptor, and this finding has led to an enormous interest into PPARy receptor biology, TZD action, and the treatment of insulin resistance. Thus, the overall goals of this research proposal are to elucidate the molecular and biochemical events in insulin'sintracellularsignalingpathway leading to GLUT4 translocation and glucose transport and to explore new mechanisms of insulin resistance. 1) We will study the post receptor signaling events which mediate insulin stimulated glucose transport. These studies are driven by our recent findings that the heterotrimeric G protein, Gaq/11, is a necessary signaling molecule in this process, conveying signals from the insulin receptor to PI3 kinase, leading to glucose transport stimulation. We will further study the role of Gaq/11 in this insulin bioeffect. 2) Insulin resistance is a major pathophysiologic feature of Type 2 diabetes mellitus, as well as a host of other conditions, and we propose novel studies of this phenomenon. Endothelin-I (ET-I) levels are elevated in Type 2 diabetic patients and other human insulin resistant states, and we have now demonstrated that treatment of 3T3-L1 adipocytes with ET-Iproduces a state of insulin resistance. We now propose a series of studies to examine the basic mechanisms of heterologous desensitization of insulin stimulated glucose transport by ET-I, as it may represent a new mechanism ofinsulin resistance in man. 3) PPARy nuclear receptors are the presumed target for anti-diabetic TZDs, and this has generated an enormous interest in the biology of this nuclear receptor. We propose detailed studies of PPARy biology, in order to help elucidate the role of this receptor, and its downstream target genes, in insulin action and insulin resistance. The proposed studies range from physiologic measurements in PPARy knockout (KO) animals, to more basic mechanistic studies of agonist mediated PPARy transcriptional events. In summary, the experiments proposed in this application will apply new methodologies to provide mechanistic information and test novel hypotheses, which will further our understanding of insulin action and insulin resistance.